GROWTH RESPONSES OF THE DIATOM,CYCLOTELLA CRYPTICA (BACILLARIOPHYCEAE), TO GIBBERELLIC ACID1

The plant hormone, gibberellic acid (GA), stimulated growth of a marine diatom, Cyclotella cryptica Reimann, Lewin and Guillard. Four concentrations of GA (5 × 20 × 25 × and 35 × 10^?6 g/mL) were added to axenic cultures of C. cryptica. Changes in cell densities, measured by cell counts and turbidimetric readings, confirmed that GA at 20 × 10^?6 g/mL produced maximum stimulation. There was an increase in the total number of cells produced and a shorter lag phase of growth at this concentration. Coulter counter measurements of cell size, as well as ocular micrometer measurements, indicated there was no significant variation in cell volumes of GA grown cells over that of the controls.     

UDPGLUCOSE PYROPHOSPHORYLASE ACTIVITY IN THE DIATOM CYCLOTELLA CRYPTICA. PATHWAY OF CHRYSOLAMINARIN BIOSYNTHESIS 1

UDPglucose pyrophosphorylase activity was detected in cell-free extracts of the diatom Cyclotella cryptica TI3L Reimann, Lewin and Guillard. When assayed in the direction of UDPglucose formation, the enzyme had maximal activity at pH 7.8 and was stimulated by Mg²⁺and Mn²⁺ions. 3-Phosphoglycerate and inorganic phosphate had little effect on enzymatic activity, and the enzyme was relatively insensitive to feedback inhibition from UDPglucose (K, > I millimolar). A glucan was formed from UDP-[¹⁴C]glucose in cell-free extracts of C. cryptica. This glucan had a median molecular weight of 4600 (as determined by gel filtration chromatograbhy) and could be hydrolyzed by laminarinase. Partial acid hydrolysis of the glucan resulted in the formation of glucose and laminaribiose. but not cellobiose. These results suggest that the synthesis of chrysolaminarin (the major storage carbohydrate of diatoms) occurs via the activity of UDPglucose pyrophosphorylase. followed by glucosyl transfer from UDPglucose to the growing β-(1–3)-linked glucan.     

AUXOSPORULATION IN CYCLOTELLA OCELLATA (BACILLARIOPHYCEAE) UNDER NATURAL AND EXPERIMENTAL CONDITIONS1

Growth and sexual reproduction in a population of Cyclotella ocellata Pantocseck were studied during one annual cycle in a reservoir and in short-term enclosure experiments performed in situ involving different nutrient conditions and concentrations of zooplankton species. Three phases of auxosporulation in this diatom were distinguishable morphologically: 1) preauxospore, from the beginning of zygote formation until the valves were longitudinally separated, 2) primary auxospore, when the zygote grew too large to fit inside the valves and before it reached its full size, and 3) mature auxospore, characterized by a well-developed, markedly scalloped edge. Under experimental and natural conditions, sexual reproduction was associated with changes in cell size. In the natural system, the auxospore appeared to act as a resting structure during conditions adverse for population growth. A threshold population of small cells appeared to be necessary for sexual reproduction in the natural system, whereas auxosporulation was associated with phosphorus fertilization in the enclosures. In both environments only cells smaller than 9.5 μm in diameter were capable of auxospore formation. Our results suggest that, once having reached the critical cell size, the factors that trigger sexual reproduction may depend on ambient environmental conditions.     

EFFECTS OF SILICON DEFICIENCY ON LIPID COMPOSITION AND METABOLISM IN THE DIATOM CYCLOTELLA CRYPTICA1

The effects of silicon deficiency on the metabolism and composition of lipids in Cyclotella cryptica T13L Reimann, Lewin, and Guillard were examined. Silicon-deficient cells had higher levels of neutral lipids (primarily triacylglycerols) and higher proportions of saturated and monounsaturated fatty acids than silicon-replete cells. After 4 h of silicon deficiency, the percentage of newly assimilated NaH¹⁴CO₃ partitioned into lipids increased from 27.6% to 54.1%, whereas the percentage partitioned into chrysolaminarin decreased from 21.6% to 10.6%. In addition, pulse-chase experiments with NaH¹⁴CO₃ indicated that the amount of ¹⁴C in the total cellular lipid fraction increased by 32% after 12 h of silicon deficiency despite the absence of additional photoassimilable ¹⁴C. Therefore, the accumulation of lipids in response to silicon deficiency appears to be due to two distinct processes: (a) an increase in the proportion of newly assimilated carbon partioned into lipids, and (2) a slow conversion of previously assimilated carbon from non-lipid compounds into lipids     

ULTRASTRUCTURE OF THE FRUSTULE OF TRICERATIUM FAVUS (BACILLARIOPHYCEAE)1,2

New structural details of the frustules of the diatom Triceratium favus Ehrenberg seen in the scanning electron microscope are reported. Significant new observations concern the pores of the hexagonal chambers, accessory structures (spines, dendritic processes) on the outer surfaces of the hexagonal chambers, the value margins and girdle structure.     

FURTHER OBSERVATIONS AND SOME COMMENTS ON THE FINE STRUCTURE OF THE CENTRIC DIATOM AULACOSEIRA ISLANDICA (BACILLARIOPHYCEAE)1

Using light and electron microscopy, the diatom species Aulacoseira islandica (O. Müll.) Sim. was examined with special emphasis on the following characteristics: structure of the valve areolae, heterovalvy, and distribution of the rimoportulae. The mantle and valve face areolae were pores containing volate occlusions. However, observations only using transmission electron microscopy may result in an incomplete interpretation because of the fragility of the dissected system of volae. Relief valves with a stepped mantle and intaglio valves with a plain mantle occurred. Another form of heterovalvy resulted from the formation of separation valves. Linking valves had spatulate spines while separation valves bore tapering spines. In Aulacoseira, the rimoportulae usually occurred near the “Ringleiste.” The presence of several rimoportulae on the mantle was one of the most striking features in Aulacoseira islandica.     

EFFECTS OF SELENIUM DEFICIENCY ON THE MORPHOLOGY AND ULTRASTRUCTURE OF THE COASTAL MARINE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)1,2

The effects of selenium deficiency on the siliceous and nonsiliceous components of the planktonic marine diatom Thalassiosira pseudonana (Hust.) Hasle and Heimdal (clone 3H) are examined using light and electron microscopy. Selenium deficiency induces elongation along the pervalvar axis initially as a result of chain formation caused by the failure of sibling cells to separate and subsequently by cell elongation via the production of hyaline girdle bands. In Se-deficient cultures cell elongation involves the blockage of both mitotic and cytokinetic components of cell division. Selenium deficiency results in ultra-structural alterations in the reticular membrane system and in mitochondrial and chloroplast membranes. Various types of inclusions are seen in vacuolar areas and the accumulation of lipid reserves is evident in Se-deficient cells. These results provide indirect evidence for a metabolic Se requirement in this algal species.     

EFFECTS OF SELENIUM DEFICIENCY ON THE MORPHOLOGY AND ULTRASTRUCTURE OF THE COASTAL MARINE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)1,2

The effects of selenium deficiency on the siliceous and nonsiliceous components of the planktonic marine diatom Thalassiosira pseudonana (Hust.) Hasle and Heimdal (clone 3H) are examined using light and electron microscopy. Selenium deficiency induces elongation along the pervalvar axis initially as a result of chain formation caused by the failure of sibling cells to separate and subsequently by cell elongation via the production of hyaline girdle bands. In Se-deficient cultures cell elongation involves the blockage of both mitotic and cytokinetic components of cell division. Selenium deficiency results in ultrastructural alterations in the reticular membrane system and in mitochondrial and chloroplast membranes. Various types of inclusions are seen in vacuolar areas and the accumulation of lipid reserves is evident in Se-deficient cells. These results provide indirect evidence for a metabolic Se requirement in this algal species.     

OBSERVATIONS OF A TUBE-DWELLING DIATOM: NAVICULA HAMULIFERA (BACILLARIOPHYCEAE)1

Colonies of the tube-dwelling diatom Navicula hamulifera Grunow living on mangrove prop roots in Indian River, Florida and at La Parguera, Puerto Rico, were studied using light and electron microscopy. Observations of the tube morphology and cell structure of this diatom from fresh samples and cultures are described, as well as the ultrastructural morphology of its frustule. The formation of tubes by this diatom is reported for the first time. Comparisons are made with the closest species; Navicula delognei V.H. and Navicula pseudoco moides Hendey.     

ORGANIZATION OF THE CHLOROPLAST GENOME OF THE FRESHWATER CENTRIC DIATOM CYCLOTELLA MENEGHINIANA1

We constructed a complete physical map and a partial gene map of the chloroplast genome of Cyclotella meneghiniana Kützing clone 1020-1a (Bacillariophyceae). The 128-kb circular molecule contains a 17-kb inverted repeat, which divides the genome into single copy regions of65 kb and 29 kb. This is the largest genome and inverted repeat found in any diatom examined to date. In addition to the 16S and 23S ribosomal RNA genes, the inverted repeat contains both the ndhD gene (as yet unexamined in other diatoms) and the psbA gene (located similarly in one of two other examined diatoms). The Cyclotella chloroplast genome exists as two equimolar populations of inversion isomers that differ in the relative orientation of their single copy sequences. This inversion heterogeneity presumably results from intramolecular recombination within the inverted repeat. For the first time, we map the ndhD, psaC, rpofi, rpoCl, and rpoC2 genes to the chloroplast genome of a chlorophyll c-containing alga. While the Cyclotella chloroplast genome retains some prokaryotic and land plant gene clusters and operons, it contains a highly rearranged gene order in the large and small single copy regions compared to all other examined diatom, algal, and land plant chloroplast genomes.     

OBSERVATIONS OF A TUBE-DWELLING DIATOM: NAVICULA HAMULIFERA (BACILLARIOPHYCEAE)1

Colonies of the tube-dwelling diatom Navicula hamulifera Grunow living on mangrove prop roots in Indian River. Florida and at La Parguera, Puerto Rico, were studied using light and electron microscopy. Observations of the tube morphology and cell structure of this diatom from fresh samples and cultures are described, as well as the ultrastructural morphology of its frustule. The formation of tubes by this diatom is reported for the first time. Comparisons are made with the closest species; Navicula delognei V.H. and Navicula pseudocomoides Hendey.     

CARBONIC ANHYDRASE IN THE MARINE DIATOM THALASSIOSIRA WEISSFLOGII (BACILLARIOPHYCEAE)1

The zinc metalloenzyme carbonic anhydrase plays a critical role in inorganic carbon acquisition in marine diatoms, thus conferring on zinc a key role in oceanic carbon cycling. As a first step in determining the location and function of carbonic anhydrase (CA) in Bacillariophyceae, we purified and partially sequenced CA from T. weissflogii (Gru) Fryxell et Hasle (TWCA1) and cloned the corresponding cDNA (twca1). The twca1 sequence is different from other known algal carbonic anhydrase genes, and encodes a protein of roughly 34 kDa. The amino terminal amino acids sequenced from purified TWCA1 are 72 residues downstream of the putative starting methionine predicted by twca1. This difference may be due to the presence of a short-lived signal sequence designed to guide the enzyme to the correct cellular location. The absence of any homology between TWCA1 and previously sequenced CAs from Chlorophyceae may indicate either convergent evolution or that carbon acquisition represents a fundamental physiological difference among algal phyla.     

COPPER SORPTION AND RELEASE BY CYCLOTELLA MENEGHINIANA (BACILLARIOPHYCEAE) AND CHLAMYDOMONAS REINHARDTII (CHLOROPHYCEAE)1

Initial Cu⁺⁺ sorption by Cyclotella meneghiniana Kütz. (Cu⁺⁺-sensitive) and Chlamydomonas reinhardtii Dangeard (Cu⁺⁺-resistant) was rapid in the first 5 min of Cu⁺⁺ incubation with little sorption after 2 h. On a cell to cell basis, Cyclotella sorbed ca. five times more Cu⁺⁺ from the medium than Chlamydomonas. In MBL medium with EDTA Cyclotella and Chlamydomonas cells sorbed 21.0 and 4.41 nM Cu⁺⁺/10⁶ cells respectively in 6 h with 0.3 mg Cu⁺⁺/l in the medium. Proportionally similar quantities of Cu⁺⁺ were sorbed when the cells were Cu⁺⁺ incubated in MBL + citrate or filtered lake water. Cleaned cell walls of Cyclotella sorbed little Cu⁺⁺ (1.7 nM/10⁶ cells) as compared to living cells (17.5 nM Cu⁺⁺/10⁶ cells) in 3 h. Therefore, in living Cyclotella most of the Cu⁺⁺ taken up must be absorbed by the protoplasm or perhaps by the organic layer surrounding the silica wall. Cleaned cell walls of Chlamydomonas sorbed 3.5 nM Cu⁺⁺/10⁶ cells and living Chlamydomonas cells sorbed 2.6 nM Cu⁺⁺/10⁶ cells. This indicates that most of the Cu⁺⁺ sorbed by Chlamydomonas cells remained bound to the cell wall and probably did not readily enter into the protoplasm: When placed in Cu⁺⁺ free medium after Cu⁺⁺ incubation, Cyclotella and Chlamydomonas cells released 46 and 59% respectively of the Cu⁺⁺ sorbed.     

PHYSIOLOGICAL AND ULTRASTRUCTURAL RESPONSES OF CYCLOTELLA MENEGHINIANA (BACILLARIOPHYTA) TO LIGHT INTENSITY AND NUTRIENT LIMITATION1

Photosynthetic characteristics and chloroplast ultrastructure of Cyclotella meneghiniana Kütz. were quantified while the organism was simultaneously adjusting to light and nutrient stress. Cells were grown in batch culture at either low or high light intensity on medium with a nitrogen/phosphorus molar ratio of 2:1 as a control, or with nitrogen or phosphorus deleted from the medium to create nutrient deficiencies. Analysis of variance indicated that light intensity, nutrient deficiency and duration of nutrient deficiency all had significant effects on cell growth, chlorophyll (Chl) concentration/cell, cellular fluorescence capacity (CFC), chloroplast volume and thylakoid surface density. Because interactions existed among nutrient deficiency, extent of nutrient deficiency, and light intensity, all three must be considered together in order to describe accurately the physiology and chloroplast ultrastructure of the diatom. Significant correlations were found between the Chl/cell or CFC/cell and chloroplast volume and thylakoid surface density. Through an increase in Chi concentration, chloroplast volume and thylakoid surface density, the cells successfully adapted to the conditions of low light intensity even while under nutrient stress. In contrast, less Chl/cell, smaller chloroplast volume and less thylakoid surface density were found at high light intensity.     

COMPARATIVE ULTRASTRUCTURE OF THREE SPECIES OF CHLOROSARCINA (CHLOROSARCINACEAE,CHLOROPHYTA)1,2

Ultrastructural studies revealed that Chlorosarcina stigmatica Deason differs from C. brevispinosa and C. longispinosa Chantanachat and Bold because the vegetative cells possess a pyrenoid, and the zoospores have chlorophycean characteristics. Zoospores of the latter species exhibit pleurastrophycean characteristics including counterclockwise absolute orientation of the flagellar apparatus components. Nuclear division in C. longispinosa, like that of Friedmannia israelensis Chantanachat and Bold, is metacentric. It is suggested that a new genus, Desmotetra, be erected for Chlorosarcina stigmatica, and that it, along with Chlorosarcinopsis Herndon, be placed in the Chlorococcales, Chlorophyceae. Chlorosarcina brevispinosa and C. longispinosa belong in the Pleurastrales, Pleurastrophyceae.     

COMPARATIVE ULTRASTRUCTURE OF THREE SPECIES OF CHLOROSARCINA (CHLOROSARCINACEAE,CHLOROPHYTA) 1,2

Ultrastructural studies revealed that Chlorosarcina stigmatica Deason differs from C. brevispinosa and C. longispinosa Chantanachat and Bold because the vegetative cell possess a pyrenoid, and the zoospores have chlorophycean characteristics. Zoospores of the latter species exhibit pleurastrophycean characteristics including counterclockwise absolute orientation of the flagellar apparatus components. Nuclear division in C. longispinosa, like that of Friedmannia israelensis Chantanachat and Bold, is metacentric. It is suggested that a new genus, Desmotetra, be erected for Chlorosarcina stigmatica, and that it, along with Chlorosarcinopsis Herndon, be placed in the Chlorococcales, Chlorophyceae. Chlorosarcina brevispinosa and C. longispinosa belong in the Pleurastrales, Pleurastrophyceae.     

OBSERVATIONS ON CHAETOCEROS BUCEROS (BACILLARIOPHYCEAE), A RARE TROPICAL PLANKTONIC SPECIES COLLECTED FROM THE MEXICAN PACIFIC1

Examination of net plankton samples from the Gulf of Tehuantepec (Mexican Pacific) yielded a rare tropical species of the genus Chaetoceros, C. buceros Karsten. This species is conspicuous because of its relatively large size, type of chain formation, and shape of the terminal setae. Electron microscopic study revealed other interesting characteristics: the intercalary values are more lightly silicified than the terminal valves and show a pattern of costae radiating form the center with various annuli (“central hyaline fields”) in the center of the valve face; both kind of valves are perforated by small poroids. Terminal valves possess several (21–30) slitlike and randomly oriented rimoportulae. Important morphological differences exist between the intercalary and the terminal setae. Morphological characters, comparision with related species, and the systematic position are discussed. A new section of the subgenus Hyalochaete, Conspicua, is proposed to include C. buceros.     

THE ULTRASTRUCTURE AND CYTOCHEMISTRY OF RESTING CELL FORMATION IN AMPHORA COFFAEFORMIS (BACILLARIOPHYCEAE)1

The ultrastructure of logarithmic-growing cells and of resting cells in laboratory cultures of Amphora coffaeformis (Ag.) Kütz. isolated from deep ocean water was examined using electron and light microscopy. The acid Phosphatase activity, chlorophyll a and lipid content were assessed at weekly intervals of resting cell formation during cold-dark treatment, simulating deep ocean water. Approximately 4 wk are required to complete resting cell formation. During the first week, the cytoplasm undergoes extensive transformation and lysosomal activity is observed. Large vacules decrease in size and many small ones develop, the mitochondria become fewer and one or more massive mitochondria appear possibly by fusion of smaller ones; the cytoplasm becomes densely granular. During the second and third week, the cytoplasm continues to contract, lipid bodies begin to develop and the plastid becomes densely stained. At the fourth week, the mature resting cell is formed containing one or more massive mitochondria, a well-formed plastid, and granular cytoplasm containing occasional lipid droplets. There is no change in frustule morphology and the cytoplasm does not produce a protective layer. The variation in chemical constituents correlates with microscopic structure of the cells. The fine structure of cells during growth resumption when exposed to light at 25 c is presented. Previous reports of viable, chlorophyll-containing cells at great depths in the ocean may be explained by the results reported in this paper.     

TWO DISTINCT COLONIAL MORPHOTYPES OF AMPHORA COFFEAEFORMIS (BACILLARIOPHYCEAE) CULTURED ON SOLID MEDIA1

When cultured on different types of solid media, the marine-fouling diatom Amphora coffeaeformis (Ag.) Kütz. consistently formed two distinct colonial morphotypes named tight and fuzzy. Tight colonies were comprised mainly of small, morphologically distorted, nonmotile cells, whereas morphologically normal and highly motile cells formed the fuzzy colonies. Cells from tight colonies were less adherent to glass, grew more slowly in liquid media, and had a slightly decreased viability on plates with copper than cells from fuzzy colonies. Whereas the protein profiles of the two types of cells were nearly identical in polyacrylamide gels stained with Coomassie blue, cells from tight colonies produced a significantly lower amount of a protease-resistant, low M_r polysaccharide or glycoconjugate as detected in silver-stained gels. The frequency of appearance of the fuzzy and tight morphotypes was not influenced by the mode of nutrition or the type of substratum to which the algal cells adhered. However, certain formulations of solid medium and the presence of growth-inhibitory concentrations of copper in agar plates favored the formation of tight colonies. Due to their frequencies and patterns of appearance, it was clear that the two naturally formed morphotypes were not the consequence of spontaneous mutations, genetic rearrangement, or selection of stable natural variants, and we have hypothesized that they were linked to a normal physiological behavior. The tight colonial morphotype was used as a valuable marker to screen for true motility/adhesion mutants within an ultraviolet-mutagenized population of A. coffeaeformis. Seven mutants were isolated that were non-motile on agar plates, poorly adherent to glass, and distinguished from naturally formed cells from tight colonies by their inability to form fuzzy colonies upon subculture on solid media.     

EFFECT OF SILICATE LIMITATION ON VALVE MORPHOLOGY IN THALASSIOSIRA AND COSCINODISCUS (BACILLARIOPHYCEAE)1

Coscinodiscus radiatus Ehrenb. and Thalassiosira eccentrica (Ehrenb.) Cleve were grown in a silicate-limited chemostat at silicate concentrations below 1 μg-atoms · l^?1. The resulting abnormal valves of C. radiatus lacked a thickened ring around the foramina; their pore membranes were thinner and their loculi shallower than those in normal cells. Abnormal valves of T. eccentrica had a fasciculate areolae pattern; they lacked a silica covering over the foramina and some tangential areolae walls. Neither abnormal valve could be termed a new species.